Plant defense against pathogens differs in its mechanism from that observed in animals. For example, there is known in higher plants a hypersensitive response (HR) mechanism which involves a dynamic resistance reaction to pathogen invasion. When a pathogen invades a plant, plant cells at a site of invasion die in response, whereby pathogens are trapped locally. This reaction is known to be induced as a result of either an incompatible host-pathogen interaction or a non-host-pathogen interaction. Such cell suicide can be understood in terms of a localized, programmed cell death (Dangl et al.: Plant Cell 8: 1973-1807 (1996)). In addition to the mechanism involving HR, other defense reactions, including generation of active oxygen species, reinforcement of a cell wall, production of phytoalexin and biosynthesis of defense-related proteins such as PR proteins, are also known (Hammond-Kosack and Jones: Plant Cell 8: 1773-1791 (1996)). Further, in addition to such localized defense responses, there is known to take place in many cases a defense reaction spreads whereby PR proteins accumulate also in non-infected parts of a plant, whereby resistance is imparted to the entire plant. This mechanism is referred to as systemic acquired resistance (SAR) and continues for several weeks or longer. As a result, the entire plant is made resistant to secondary infection (Sticher et al.: Annu. Rev. Phytopathol. 35: 235-270 (1997)).
A first reaction of a plant of switching on a highly organized defense reaction such as outlined above is the recognition by the plant of a molecule called an “elicitor” directly or indirectly produced by an invading pathogen. Additionally, complex signal cascades including the subsequent rapid generation of active oxygen species and reversible protein phosphorylation are considered to be important as initial reactions of the defense response (Yang et al.: Genes Dev. 11: 1621-1639 (1997)). There are a wide variety of elicitors, including so-called non-specific elicitors e.g. oligosaccharides which are products by degradation of cell wall components of many fungi including chitin/chitosan and glucan, or oligogalacturonic acids derived from a plant cell wall, variety-specific elicitors e.g. avirulence gene products of pathogens such as AVR 9 (Avr gene products), and elicitors with an intermediate specificity such as elicitin (Boller: Annu. Rev. Plant Physiol. Plant Mol. Biol. 46: 189-214 (1995)).
Harpin is a bacterium-derived protein elicitor which induces hypersensitive cell death in a non-host plant (Wei et al.: Science 257: 85-88 (1992), He et al.: Cell 73: 1255-1266 (1993)). Harpin (harpinEa) has been purified as a first bacterium-derived HR-inducing protein from Erwinia amylovora Ea321, a pathogen of pear and apple, and Escherichia coli transformed with a cosmid containing the hrp gene cluster, and an hrpN gene encoding Harpin has been cloned (Wei et al.: Science 257: 85-88 (1992)). Thereafter, harpinpss encoded by hrpZ gene has been identified and characterized from Pseudomonas syringae pv. syringae 61, a pathogen of a bean, by screening an Escherichia coli expression library with an activity of inducing HR to a tobacco leaf as an index (He et al.: Cell 73: 1255-1266 (1993), and Japanese Patent Application Domestic Announcement No. 1996-510127). The homology between these two harpins is low, and a relatively high homology is found only in 22 amino acids. Moreover, the role of a harpin in pathogenicity has not been made clear. In addition to these, as a third protein, PopA protein (which PopA encodes) is identified from Pseudomonas solanacearum GMI1000, a pathogen of a tomato, as a protein inducing HR to a non-host tobacco (Arlat et al.: EMBO. J. 13: 543-553 (1994)). Though PopA gene is located on the outside of hrp cluster, differing from hrpN and hrpZ, they are identical in that they are under the control of an hrp regulon. The above three proteins are glycine-rich, heat stable proteins, induce HR to a non-host tobacco and are secreted extracellularly at least in vitro in a manner of depending upon hrp protein. In addition to these are reported HrpW protein from Pseudomonas syringaepv. tomato DC3000 as a protein having the same function (Charkowski et al.: J. Bacteriol. 180: 5211-5217 (1998)), hrpZpst and hrpZpsg proteins as harpinpss homologues (Preston et al.: Mol. Plant-Microbe. Interact. 8: 717-732 (1995)), and harpinEch (Bauer et al.: Mol. Plant-Microbe. Interact. 8: 484-491 (1995)) and hrpNECC protein (Cui et al.: Mol. Plant-Microbe. Interact. 9: 565-573 (1996)) as harpinEa homologues.
It has been made apparent from studies upon various metabolic inhibitors that the formation of localized necrosis spots with harpin is not so-called necrosis due to the cytotoxicity of harpin but a cell death resulting from a positive response on the plant side (He et al.: Mol. Plant-Microbe. Interact. 7: 289-292 (1994), and He et al.: Cell 73: 1255-1266 (1993)), and this hypersensitive cell death is thought to be a type of programmed cell death (Desikan et al.: Biochem. J. 330: 115-120 (1998)). The addition of harpinpss into a cell culture of Arabidopsis induces a homologue of gp91-phox, a constituent of NADPH oxidase, which is thought to have an important role in the oxidative burst as an initial reaction of a disease-resistant reaction, (J. Exp. Bot. 49: 1767-1771 (1998)), and mitogen-activated protein (MAP) kinase (Desikan et al.: Planta. 210: 97-103 (1999)). Moreover, a harpin can impart systemic acquired resistance (SAR) to a plant. For example, SAR meditated by salicylic acid and an NIM gene can be induced to an Arabidopsis plant by artificially injecting harpinEa into the plant cells (Dong et al.: The Plant J. 20: 207-215 (1999)), and Harpinpss can induce SAR to a cucumber and impart a wide spectrum of resistance to fungi, viruses and bacteria (Strobel et al.: Plant J. 9: 431-439 (1996)).
Thus, there are reports about artificially injecting or spraying purified harpin into a plant and analyzing the induction of a hypersensitive cell death and an acquired resistance reaction (Japanese Patent Application Domestic Announcement No. 1999-506938, Strobel et al.: Plant J. 9: 431-439 (1996), and Dong et al.: The Plant J. 20: 207-215 (1999)). However, there is no report about introducing a gene encoding an elicitor protein such as a harpin into a plant to produce a transgenic plant and analyzing it.